Environmental stress due to salinity is one of the most serious factors limiting the productivity of agricultural crops, which are predominantly sensitive to the presence of high concentrations of salts in the soil. Large terrestrial areas of the world are affected by levels of salt inimical to plant growth. It is estimated that 35-45% of the 279 million hectares of land under irrigation is presently affected by salinity. This is exclusive of the regions classified as arid and desert lands, (which comprises 25% of the total land of our planet). Salinity has been an important factor in human history and in the life spans of agricultural systems. Salt impinging on agricultural soils has created instability and has frequently destroyed ancient and recent agrarian societies. The Sumerian culture faded as a power in the ancient world due to salt accumulation in the valleys of the Euphrates and Tigris rivers. Large areas of the Indian subcontinent have been rendered unproductive through salt accumulation and poor irrigation practices. In this century, other areas, including vast regions of Australia, Europe, southwest USA, the Canadian prairies and others have seen considerable declines in crop productivity.
Although there is engineering technology available to combat this problem, though drainage and supply of high quality water, these measures are extremely costly. In most of the cases, due to the increased need for extensive agriculture, neither improved irrigation efficiency nor the installation of drainage systems is applicable. Moreover, in the arid and semi-arid regions of the world water evaporation exceeds precipitation. These soils are inherently high in salt and require vast amounts of irrigation to become productive. Since irrigation water contains dissolved salts and minerals, an application of water is also an application of salt that compounds the salinity problem.
Increasing emphasis is being given to modify plants to fit the restrictive growing conditions imposed by salinity and even bioremediate the soil through extraction of the salt. If economically important crops could be manipulated and made salt resistant, this land could be farmed resulting in greater sales of seed and greater yield of useful crops. Conventional breeding for salt tolerance has been attempted for a long time. These breeding practices have been based mainly on the following strategies: a) the use of wide crosses between crop plants and their more salt-tolerant wild relatives, b) screening and selecting for variation within a particular phenotype, c) designing new phenotypes through recurrent selection. (Rush, et al. (1981); Norlyn (1980) and Tal (1985) The lack of success in generating tolerant varieties (given the low number of varieties released and their limited salt tolerance) would suggest that conventional breeding practices are not enough and that in order to succeed a breeding program should include the engineering of transgenic crops. (Flowers, et al. (1995) and Bonhert, et al. (1996))
Several biochemical pathways associated with stress tolerance have been characterized in different plants and a few of the genes involved in these processes have been identified and in some cases the possible role of proteins has been investigated in transgenic/overexpression experiments. Several compatible solutes have been proposed to play a role in osmoregulation under stress. Such compatible solutes, including carbohydrates, amino acids and quaternary N-compounds have been shown to increase osmoregulation under stress. (Tarcynski, et al. (1995); Kishor (1995) and Ishitani (1995)) Also, proteins that are normally expressed during seed maturation (LEAs, Late Embryogenesis Abundant proteins) have been suggested to play a role in water retention and in the protection of other proteins during stress. The overexpression of LEA in rice provided a moderate benefit to the plants during water stress. (Xu, et al. (1996) and Wu, et al. PCT # WO/9713843) A single gene (sod2) coding for a Na+/H+ antiport has been shown to confer sodium tolerance in fission yeast. (Jia, et al. (1992) and Young, et al., PCT # WO/0106651) One of the main disadvantages of using this gene for transformation of plants is associated with the typical problems encountered in heterologous gene expression, i.e. incorrect folding of the gene product, targeting of the protein to the target membrane and regulation of the protein function.
Plants that tolerate and grow in saline environments have high intracellular salt levels. A major component of the osmotic adjustment in these cells is accomplished by ion uptake. The utilization of inorganic ions for osmotic adjustment suggests that salt-tolerant plants must be able to tolerate high levels of salts within their cells. However, enzymes extracted from these plants show high sensitivity to salt. The sensitivity of the cytosolic enzymes to salt would suggest that the maintenance of low cytosolic sodium concentration, either by compartmentation in cell organelles or by exclusion through the plasma membrane, must be necessary if the enzymes in the cell are to be protected from the inimical effects of salt.
Plant cells are structurally well suited to the compartmentation of ions. Large membrane-bound vacuoles are the site for a considerable amount of sequestration of ions and other osmotically active substances. A comparison of ion distribution in cells and tissues of various plant species indicates that a primary characteristic of salt tolerant plants is their ability to exclude sodium out of the cell and to take up sodium and to sequester it in the cell vacuoles. Transport mechanisms could actively move ions into the vacuole, removing the potentially harmful ions from the cytosol. These ions, in turn, could act as an osmoticum within the vacuole, which would then be responsible for maintaining water flow into the cell. Thus, at the cellular level both specific transport systems for sodium accumulation in the vacuole and sodium extrusion out of the cell are correlated with salt tolerance. It would be a particular advantage to use plants that accumulate salt in the vacuole in response to high salt in the soil. Such plants would accumulate the salt in the leaves and roots, which can be removed, removing a portion of the salt.
Furthermore, profits in the cattle industry are affected by the high cost of labor; thus, management procedures which reduce labor requirements are important. One management tool frequently used is regulating feed intake with salt. Self-feeding supplements tend to allow timid, slow-eating cows to get their share and it is an easy method of providing Vitamin A, phosphorus and other feed additives. Because there are practical limits to the amount of salt cattle eat, salt can also be used to restrict the consumption of highly palatable feeds such as grain and supplement. Salt is also added to feed grain because with high grain rations, urinary calculi (phosphatic type) are a problem. This problem is controlled by feeding salt to flush out the stones. This is particularly true with milo and cottonseed meal based finishing programs. Salt supplements are added directly to the feed directly in the proportions desired. The addition and mixing requires labor which reduces profits. Thus there is a need for feed plants that already contain salt.
There is a long felt need in the art for the in situ remediation of soils damaged by accumulation of salts. The present invention enables phytoremediation and/or revegetation of contaminated environments via salt tolerant plants. The plants of the present invention may be grown in high salt soil and will accumulate salt in the leaves and roots. Such salt containing plant materials may be used as forage for cattle with the additional advantage that extra salt need not be added to the feed.